Dr. Barr has the long-term goal of pursuing independent investigation in the field of somatic gene therapy. Receipt of a Clinical Investigator Award will facilitate the development of Dr. Barr's investigative skills and experience by expanding his knowledge of several new cellular techniques as outlined in the proposed studies. Dr. Barr and his sponsor, Dr. Jeffrey M. Leiden, have developed an educational environment which will provide the candidate essential training in adenovirus biology and immunology. Advisors have been identified to mentor Dr. Barr's progression to an independent investigator. The proposed application will permit the candidate to develop a career as an academic physician- scientist committed to basic medical research into somatic gene therapy approaches to the treatment of inherited and acquired diseases. Over the next 5 years, Dr. Barr will focus on developing such approaches for the treatment of serum protein deficiencies. Currently, a number of these diseases are treated with repeated subcutaneous or intravenous infusions of purified or recombinant proteins. These include hemophilias A and B, treated with purified factors VIII or IX, diabetes mellitus, treated with insulin, and the erythropoietin-responsive anemias, treated with recombinant human erythropoietin (hEpo). Because each of these diseases requires life-long therapy, the development of a practical in vivo gene transfer approach for the stable delivery of physiologically significant levels of recombinant proteins to the systemic circulation would represent a significant advance in the treatment of these diseases. The overall goal of the studies outlined in this proposal is to develop such a system utilizing intramuscular (IM) injections of replication-defective recombinant adenoviruses. The method to be used is based upon recent observations made by Dr. Barr in Dr. Leiden's laboratory that (i) genetically modified human myoblasts can be used to produce stable and physiologically significant levels of recombinant serum proteins in the circulation following IM injection, (ii) replication-defective adenoviruses can program stable recombinant gene expression in a large proportion of skeletal myocytes following IM injection of neonatal or immunocompromised mice, and (iii) IM injection of such vectors into adult immunocompetent animals results in efficient but transient recombinant gene expression due to an immune response to the adenovirus-infected cells. In the studies described in this application, Dr. Barr will use the erythropoietin-responsive anemias as a model system to explore the feasibility of using IM injections of replication-defective adenoviruses for the treatment of acquired and inherited serum protein deficiencies. Specifically, Dr. Barr will (i) optimize the efficiency and stability of recombinant, replication-defective adenovirus-mediated in vivo gene transfer into skeletal muscle in neonatal and adult mice, (ii) assess the stability and physiologic consequences of recombinant hEpo secretion following IM injection of neonatal and adult immunocompetent and SCID mice with an adenovirus encoding the hEpo cDNA, (iii) elucidate the cellular elements underlying the immune response to adenovirus-infected cells in vivo, (iv) construct a new generation of replication-defective adenoviruses containing deletions in the E1, E3, and E4 regions that should eliminate any low-level late viral protein synthesis in vivo, and (v) test these new adenoviruses in adult immunocompetent mice. The results of these studies will be directly applicable to the treatment of the erythropoietin-responsive anemias associated with both HIV infection and chronic renal failure, diseases affecting over 150,000 Americans. In addition, they will have important implications for the treatment of many other inherited or acquired serum protein deficiencies.